Alejandro F. Schinder - US grants

DESCRIPTION (provided by applicant): Age is the major risk factor for neurodegenerative diseases. As society age's neurodegeneration become increasingly common, yet the molecular and cellular mechanisms underlying this complex condition remain largely unknown. The goal of this proposal is to explore novel avenues to investigate mechanisms of neurodegeneration in an unbiased manner. A misexpression screen that takes advantage of the power of Drosophila genetics is used to identify genes that may be involved in late-onset behavioral defects (aging). Novel genes that are thus identified to be involved in late neuronal dysfunction can be then characterized and validated in the Drosophila central nervous system by combining genetics, imaging and behavior. Effects of altered gene function can also be tested and validated in the mouse brain, by retrovirus-mediated expression in neurons of the young adult and aging brain. Cell-autonomous effects emerging from these studies will largely contribute to understanding the specific aspects of neuronal function that are affected by the candidate gene. In parallel, a transgenic mouse model will be built based upon the gene identified in Drosophila. Altered gene function in the mouse brain will serve as a tool to characterize the effects on neuronal networks and, ultimately, on brain function, from simple locomotor behaviors to complex learning traits. In addition, transgenic mice should in the future be amenable to high throughput chemical/ small molecule library screens to identify drugs interfering with the degenerative process. A particular gene that emerged from the fly screen, enabled, has been selected as a proof of principle for the approach. The success of the proposed project relies on building a close interaction among two laboratories at the Leloir Institute (Buenos Aires) and a laboratory at Childrens Hospital, Harvard Medical School. Graduate students, postdoctoral fellows and young investigators will have the opportunity to obtain training at Harvard to generate, characterize and maintain transgenic mice. This know-how will certainly improve the capabilities to develop novel models of neurodegeneration at the foreign institution. The ultimate goal is to establish a long-term collaboration in order to bring our local scientific community in closer contact with novel technologies for research on this challenging topic. To accomplish such goal the preparation of full RO1 research proposal is warranted. PUBLIC HEALTH RELEVANCE: Age is a major risk factor for neurodegenerative diseases, which cause a terrible human toll. As society ages, neurodegenerative diseases will become increasingly common, underscoring the need for an in depth understanding of the molecular mechanisms underlying these devastating diseases, as well as the development of specific treatments.

DESCRIPTION (provided by applicant): The adult dentate gyrus continuously generates granule cells (DGCs) that are needed for specific learning and memory tasks, but the precise contribution of new neurons to information processing in the hippocampal circuitry remains unknown. In the past, we have demonstrated that key developmental processes occurring in the perinatal brain such as maturation of excitability, afferent synaptogenesis and function are all recapitulated during adult neurogenesis. We now propose the central hypothesis that newly born cells establish functional outputs as they develop, and the population of postsynaptic target cells contacted by young neurons is predominantly inhibitory, therefore different from the mixed excitatory/inhibitory network activated by mature granule cells. Thus, there would be a time window in which young DGCs primarily activate feedforward and/or feedback inhibitory circuits without exciting pyramidal cells, exerting a tight inhibitory control over the dentate gyrus output. In Aim 1 we will build a spatio-temporal map of target activation by young developing neurons of the adult dentate gyrus. We will use retroviral transduction to express the light- activated cation channel Channelrhodopsin-2 in newborn DGCs of young-adult female mice (C57Bl6/J). We will sacrifice the animals at different times and prepare acute brain slices to carry out electrophysiological recordings. By stimulating the whole hippocampal slice with brief light pulses, all retrovirally transduced neurons will spike. We will search randomly for active postsynaptic target cells throughout the hilus and CA3 regions, and identify and characterize responsive neurons by combining loose patch and whole-cell recordings. In Aim 2 we will investigate the functional maturation of new mossy fiber synapses made onto GABAergic and glutamatergic targets. We plan to utilize whole-cell recordings to test whether synapses forming onto GABAergic interneuron targets mature faster than those made onto pyramidal cells, as suggested by our previous structural studies. We will also study presynaptic mechanisms of short- and long-term plasticity that will shape both activity-dependent competition and activation of postsynaptic circuits. This project will address fundamental questions about connectivity and activation (spiking) of newborn cells that will contribute to understanding the precise impact of adult neurogenesis in the preexisting hippocampal network and the rules of neuronal connectivity in the adult brain. Identifying the rules by which neurons integrate in the existing network in a manner that is both safe and functionally relevant is also crucial for developing future brain repair therapies. Novel retroviral tools will be developed in collaboration with the Gage lab to enhance Channelrhodopsin-2 expression, thus improving the capability of light activation of newborn cells. In turn, experimental data obtained here will be used to feed into the theoretical model being developed by the Gage lab on the role of immature neurons in signal processing. The success of the proposed project relies on strengthening the close interaction between the Schinder laboratory at the Leloir Institute (Buenos Aires) and the Gage laboratory at the Salk Institute of La Jolla. Members of the Schinder lab will have the opportunity to train at Salk on the generation and characterization of novel retroviral vectors, improving the capabilities to develop advanced molecular tools at the foreign institution. We anticipate that capacity building leading to the design, generation and use of novel retroviral tools at the Leloir Institute will have an enormous impact on the local scientific community. This collaborative effort will therefore serve as a driving force to increase the critical mass of Argentine investigators that incorporate competitive technologies for the study of neurodegenerative disorders within their research focus. An increase in the number of laboratories carrying out regeneration- related projects will certainly enhance awareness to these and related problems to our community. PUBLIC HEALTH RELEVANCE: There is a lot of expectation in our society on the potential of neural progenitor cells as powerful tools for brain repair, and we will need to understand how adult-born neurons establish their connectivity in the adult healthy brain before attempts can be made in case of neurodegeneration or trauma. Solid research on functional integration of newborn neurons of the adult hippocampus will certainly contribute to the development of regenerative therapies for the treatment of disorders occurring in non-neurogenic areas. In particular, it is critical to identify the rules by which neurons integrate in the adult brain circuits in a manner that is both safe and functionally relevant. This will be a major outcome of the proposed project.